Two novel related yeast nucleoporins Nup170p and Nup157p: complementation with the vertebrate homologue Nup155p and functional interactions with the yeast nuclear pore-membrane protein Pom152p.

Abstract

Abstract. We have taken a combined genetic and biochemical approach to identify major constituents of the yeast nuclear pore complex (NPC). A synthetic lethal screen was used to identify proteins which interact genetically with the major pore-membrane protein Pom152p. In parallel, polypeptides present in similar amounts to Pom152p in a highly enriched preparation of yeast NPCs have been characterized by direct microsequencing. These approaches have led to the identification of two novel and major nucleoporins, Nupl70p and Nup157p. Both Nupl70p and Nup157p are similar to each other and to an abundant mammalian nucleoporin, Nup155p (Radu, A., G. Blobel, and R. W. Wozniak. 1993. J. Cell Biol. 121: 1-9) and interestingly, nup170 mutants can be complemented with mammalian NUP155. In addition, the synthetic lethal screen identified genetic interactions between Pom152p and two other major nucleoporins, Nup188p (Nehrbass, U., S. Maguire, M. Rout, G. Blobel, and R. W. Wozniak, manuscript submitted for publication), and Nic96p (Grandi, P., V. Doye, and E. C. Hurt. 1993.EMBO J. 12: 3061-71). We have determined that together, Nup170p, Nup157p, Pom152p, Nup188p, and Nic96p comprise greater than one-fifth of the mass of the isolated yeast NPC. Examination of the genetic interactions between these proteins indicate that while deletion of either POM152, NUP170, or NUP188 alone is not lethal, pairwise combinations are. Deletion of NUP157 is also not lethal. However, nup157 null mutants, while lethal in combination with nup170 and nup188 null alleles, are not synthetically lethal with pom152 null alleles. We suggest that Nupl70p and Nup157p may be part of a morphologically symmetrical but functionally distinct substructure of the yeast NPC, e.g., the nucleoplasmic and cytoplasmic rings. Finally, we observed morphological abnormalities in the nuclear envelope as a function of alterations in the expression levels of NUP170 suggesting a specific stoichiometric relationship between NPC components is required for the maintenance of normal nuclear structure. COMMUNICATION between the cytoplasm and the nucleoplasm is controlled by nuclear pore complexes (NPCs) 1 which extend across the nuclear envelope (NE) at circular openings along the surface of the nucleus. The NPCs are tightly associated with the NE via the pore membrane domain, a specialized subdomain of the NE which lines the pore and connects the outer and inner nuclear membranes. A major function of the NPC is to regulate the bidirectional transport of proteins and ribonuAddress correspondence to Richard W. Wozniak, Department of Anatomy and Cell Biology, 5-14 Medical Sciences Building, University of Alberta, Edmonton, Alberta, Canada, T6G 2A7. Ph.: 403-492-1384. Fax: 403-492-0450. e-mail: rwozniak@anat.med.ualberta.ca. Abbreviations used in this paper:ORF, open reading frame; NE, nuclear envelope; NPC, nuclear pore complex; nups, nucleoporins; poms, pore membrane proteins; SM, synthetic minimal medium.cleoproteins between the cytoplasm and the nucleoplasm. The mechanism of mac...

GLE2, a Saccharomyces cerevisiae homologue of the Schizosaccharomyces pombe export factor RAE1, is required for nuclear pore complex structure and function.

“…Therefore, two likely complementing genes remained. One of the ORFs encoded the amino terminal portion of a known nucleoporin, Nupl57p (Aitchison et al, 1995b). However, a plasmid harboring only full-length NUP157 also did not rescue the gle2-1 Sec-and TS-phenotypes.…”

“…In many cases, additional layers of double membranes studded with densities resembling NPCs were observed in the nucleus juxtaposed below the cluster of herniated NPCs (arrow, Figure 7, (Wente and Blobel, 1993) and when NUP170 is overexpressed in poml52A cells (Aitchison et al, 1995b). The NPC clusters, herniations, and intranuclear annulate lamellae were not found in wild-type cells grown at 37°C (Figure 7D).…”

“…Such "synthetic lethal" screens with nspl, nup49, nupl, and poml52 have isolated mutated alleles of at least 10 other NPC-associated proteins required for NPC assembly and/or transport (Wimmer et al, 1992;Belanger et al, 1994;Aitchison et al, 1995b;Doye and Hurt, 1995). In this study, a synthetic lethal genetic screen was conducted with the nuplOO null (A) mutation.…”

Integrative structure and functional anatomy of a nuclear pore complex

“…The remainder of each diagonal column is made of Nup157 and Nup170, which flank Nic96 (Fig. 5B); Nup157 and Nup170 are functionally redundant, but are synthetically lethal 15 and together form another essential element of the diagonal column. Inter-spoke connections represent a second crucial stabilizing element.…”

Vertebrate Nup53 Interacts with the Nuclear Lamina and Is Required for the Assembly of a Nup93-containing Complex

“…Genetic and biochemical analyses have established that yeast Nup53p is part of a network of interacting nups that includes Nup170p, Nup59p, Nic96p, Nup157p, and Nup192p (Aitchison et al, 1995b;Marelli et al, 1998;Kosova et al, 1999;Fahrenkrog et al, 2000;Lusk et al, 2002;Makhnevych et al, 2003). A model for how these nups are organized within the NPC is beginning to emerge.…”